RESUMO
BACKGROUND: The number of recognised distinct autoimmune diseases (AIDs) has progressively increased over the years with more than 100 being reported today. The natural history of AIDs is characterized by progression from latent and subclinical to clinical stages and is associated with the presence of the specific circulating autoantibodies. Once presented, AIDs are generally chronic conditions. AIDs have the tendency to cluster and co-occur in a single patient. Autoimmune thyroid diseases (AITD) are the most prevalent of AIDs in the world population, and about one-third of the AITD patients also present with a non-thyroid AID during their life-span. Furthermore, patient with non-thyroid AIDs often presents with a form of AITD as a concurrent condition. Many of the clusters of AIDs are well characterized as distinctive syndromes, while some are infrequent and only described in case reports. PURPOSE: In this review, we describe the wide spectrum of the combinations and the intricate relationships between AITD and the other AIDs, excluding Addison's disease. These combinations are collectively termed type 3 Autoimmune Polyglandular Syndrome (APS-3), also called type 3 Multiple Autoimmune Syndrome (MAS-3), and represent the most frequent APS in the world populations. CONCLUSIONS: Numerous associations of AITD with various AIDs could be viewed as if the other AIDs were gravitating like satellites around AITD located in the center of a progressively expanding galaxy of autoimmunity.
Assuntos
Doença de Addison , Doença de Hashimoto , Poliendocrinopatias Autoimunes , Humanos , Poliendocrinopatias Autoimunes/diagnóstico , Poliendocrinopatias Autoimunes/epidemiologia , Autoanticorpos , SíndromeRESUMO
BACKGROUND: Autoimmune Polyglandular Syndrome type 1 (APS-1) is a rare recessive inherited disease, caused by AutoImmune Regulator (AIRE) gene mutations and characterized by three major manifestations: chronic mucocutaneous candidiasis (CMC), chronic hypoparathyroidism (CH) and Addison's disease (AD). METHODS: Autoimmune conditions and associated autoantibodies (Abs) were analyzed in 158 Italian patients (103 females and 55 males; F/M 1.9/1) at the onset and during a follow-up of 23.7 ± 15.1 years. AIRE mutations were determined. RESULTS: The prevalence of APS-1 was 2.6 cases/million (range 0.5-17 in different regions). At the onset 93% of patients presented with one or more components of the classical triad and 7% with other components. At the end of follow-up, 86.1% had CH, 77.2% AD, 74.7% CMC, 49.5% premature menopause, 29.7% autoimmune intestinal dysfunction, 27.8% autoimmune thyroid diseases, 25.9% autoimmune gastritis/pernicious anemia, 25.3% ectodermal dystrophy, 24% alopecia, 21.5% autoimmune hepatitis, 17% vitiligo, 13.3% cholelithiasis, 5.7% connective diseases, 4.4% asplenia, 2.5% celiac disease and 13.9% cancer. Overall, 991 diseases (6.3 diseases/patient) were found. Interferon-ω Abs (IFNωAbs) were positive in 91.1% of patients. Overall mortality was 14.6%. The AIRE mutation R139X was found in 21.3% of tested alleles, R257X in 11.8%, W78R in 11.4%, C322fsX372 in 8.8%, T16M in 6.2%, R203X in 4%, and A21V in 2.9%. Less frequent mutations were present in 12.9%, very rare in 9.6% while no mutations in 11% of the cases. CONCLUSIONS: In Italy, APS-1 is a rare disorder presenting with the three major manifestations and associated with different AIRE gene mutations. IFNωAbs are markers of APS-1 and other organ-specific autoantibodies are markers of clinical, subclinical or potential autoimmune conditions.
Assuntos
Doença de Addison , Candidíase Mucocutânea Crônica , Hipoparatireoidismo , Interferon Tipo I/imunologia , Poliendocrinopatias Autoimunes , Fatores de Transcrição/genética , Doença de Addison/diagnóstico , Doença de Addison/etiologia , Adulto , Autoanticorpos/sangue , Candidíase Mucocutânea Crônica/diagnóstico , Candidíase Mucocutânea Crônica/etiologia , Feminino , Humanos , Hipoparatireoidismo/diagnóstico , Hipoparatireoidismo/etiologia , Itália/epidemiologia , Masculino , Mortalidade , Mutação , Poliendocrinopatias Autoimunes/diagnóstico , Poliendocrinopatias Autoimunes/genética , Poliendocrinopatias Autoimunes/mortalidade , Poliendocrinopatias Autoimunes/fisiopatologia , Prevalência , Proteína AIRERESUMO
BACKGROUND: Addison's disease (AD) is a rare disorder and among adult population in developed countries is most commonly caused by autoimmunity. In contrast, in children genetic causes are responsible for AD in the majority of patients. PURPOSE: This review describes epidemiology, pathogenesis, genetics, natural history, clinical manifestations, immunological markers and diagnostic strategies in patients with AD. Standard care treatments including the management of patients during pregnancy and adrenal crises consistent with the recent consensus statement of the European Consortium and the Endocrine Society Clinical Practice Guideline are described. In addition, emerging therapies designed to improve the quality of life and new strategies to modify the natural history of autoimmune AD are discussed. CONCLUSIONS: Progress in optimizing replacement therapy for patients with AD has allowed the patients to lead a normal life. However, continuous education of patients and health care professionals of ever-present danger of adrenal crisis is essential to save lives of patients with AD.
Assuntos
Doença de Addison/diagnóstico , Doença de Addison/epidemiologia , Terapia de Reposição Hormonal , Doença de Addison/tratamento farmacológico , Doença de Addison/etiologia , Corticosteroides/uso terapêutico , Adulto , Fatores Etários , Feminino , Humanos , Incidência , Masculino , Prevalência , Qualidade de Vida , Fatores SexuaisRESUMO
The availability of human monoclonal antibodies (MAbs) to the TSHR has enabled major advances in our understanding of how TSHR autoantibodies interact with the receptor. These advances include determination of the crystal structures of the TSHR LRD in complex with a stimulating autoantibody (M22) and with a blocking type autoantibody (K1-70). The high affinity of MAbs for the TSHR makes them particularly suitable for use as ligands in assays for patient serum TSHR autoantibodies. Also, M22 and K1-70 are effective at low concentrations in vivo as TSHR agonists and antagonists respectively. K1-70 has important potential in the treatment of the hyperthyroidism of Graves' disease and Graves' ophthalmopathy. Small molecule TSHR antagonists described to date do not appear to have the potency and/or specificity shown by K1-70. New models of the TSHR ECD in complex with various ligands have been built. These models suggest that initial binding of TSH to the TSHR causes a conformational change in the hormone. This opens a positively charged pocket in receptor-bound TSH which attracts the negatively charged sulphated tyrosine 385 on the hinge region of the receptor. The ensuing movement of the receptor's hinge region may then cause activation. Similar activation mechanisms seem to take place in the case of FSH and the FSHR and LH and the LHR. However, stimulating TSHR autoantibodies do not appear to activate the TSHR in the same way as TSH.
Assuntos
Autoanticorpos/imunologia , Receptores da Tireotropina/imunologia , Animais , Anticorpos Monoclonais/imunologia , Glicosilação , Humanos , Modelos Moleculares , Receptores da Tireotropina/agonistas , Receptores da Tireotropina/antagonistas & inibidores , Bibliotecas de Moléculas Pequenas/farmacologiaRESUMO
Steroidogenic enzyme autoantibodies (SEAbs) are frequently present and are markers of autoimmune premature ovarian failure (POF) in females with autoimmune Addison's disease (AAD). The prevalence and significance of SEAbs in males with AAD have not yet been defined. We studied the prevalence of SEAbs in a large cohort of males with AAD and assessed the relationship between SEAbs positivity and testicular function. A total of 154 males with AAD (mean age 34 years) were studied. SEAbs included autoantibodies to steroid-producing cells (StCA), detected by immunofluorescence, and steroid 17α-hydroxylase (17α-OHAbs) and side chain cleavage enzyme (SCCAbs) measured by immunoprecipitation assays. Gonadal function was evaluated by measuring follicle-stimulating hormone (FSH), luteinizing hormone (LH), total testosterone (TT), sex hormone-binding globulin (SHGB), anti-müllerian hormone (AMH) and inhibin-B (I-B). Twenty-six males, 10 SEAbs((+)) and 16 SEAbs((-)), were followed-up for a mean period of 7·6 years to assess the behaviour of SEAbs and testicular function. SEAbs were found in 24·7% of males with AAD, with the highest frequency in patients with autoimmune polyendocrine syndrome type 1 (APS-1). The levels of reproductive hormones in 30 SEAbs((+)) males were in the normal range according to age and were not significantly different compared to 55 SEAbs((-)) males (P > 0·05). During follow-up, both SEAbs((+)) and SEAbs((-)) patients maintained normal testicular function. SEAbs were found with high frequency in males with AAD; however, they were not associated with testicular failure. This study suggests that the diagnostic value of SEAbs in males with AAD differs compared to females, and this may be related to the immunoprivileged status of the testis.
Assuntos
Doença de Addison/enzimologia , Doença de Addison/imunologia , Autoanticorpos/imunologia , Esteroides/metabolismo , Testículo/enzimologia , Testículo/imunologia , Doença de Addison/sangue , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Pré-Escolar , Seguimentos , Hormônios Gonadais/sangue , Humanos , Masculino , Pessoa de Meia-Idade , Testículo/metabolismo , Adulto JovemRESUMO
AIMS: The prevalence of autoantibodies to zinc transporter 8 (ZnT8) in Czech children at the onset of Type 1 diabetes mellitus and dynamic changes in ZnT8 autoantibody levels during disease progression were studied. The value of ZnT8 autoantibody measurements in diagnosis of Type 1 diabetes was assessed. METHODS: Serum samples from 227 children with newly diagnosed Type 1 diabetes and from 101 control children without diabetes were analysed in a retrospective cross-sectional study. One hundred and seventy-one samples from 116 of the patients with diabetes were analysed in a follow-up study at (median) intervals of 1, 3, 5 and 10 years after onset of Type 1 diabetes. ZnT8 autoantibodies were measured using a bridging enzyme-linked immunosorbent assay, while antibodies to glutamic acid decarboxylase, insulinoma antigen 2 and insulin were measured by radioimmunoassays. RESULTS: ZnT8 autoantibodies were detected in 163/227 (72%) of children at Type 1 diabetes onset and in 1/101 (1%) of the control subjects. Sixteen out of 227 (7%) patients with Type 1 diabetes were antibody negative based on three antibodies (glutamic acid decarboxylase, insulinoma antigen 2 and insulin). This false-negative rate was reduced to 10/227 (4.4%) (P < 0.05) after inclusion of ZnT8 autoantibody measurements. Of the children, 142/227 (63%) were positive for at least three antibodies and the most common combination was insulinoma antigen 2, glutamic acid decarboxylase and ZnT8. ZnT8 autoantibody levels decreased over time after Type 1 diabetes onset and the presence and level of ZnT8 autoantibodies correlated with IA-2 autoantibodies. CONCLUSIONS: A ZnT8 autoantibody enzyme-linked immunosorbent assay showed 72% disease sensitivity and 99% specificity at Type 1 diabetes onset. Measurements of ZnT8 autoantibodies are important for Type 1 diabetes diagnosis and should be included in the panel of autoantibodies tested at the onset of Type 1 diabetes.
Assuntos
Autoanticorpos/sangue , Proteínas de Transporte de Cátions/imunologia , Diabetes Mellitus Tipo 1/imunologia , Adolescente , Idade de Início , Estudos de Casos e Controles , Criança , Pré-Escolar , Estudos Transversais , República Tcheca/epidemiologia , Diabetes Mellitus Tipo 1/sangue , Diabetes Mellitus Tipo 1/epidemiologia , Humanos , Lactente , Estudos Soroepidemiológicos , Fatores de Tempo , Adulto Jovem , Transportador 8 de ZincoRESUMO
CONTEXT: Sequential conversion of Hashimoto's thyroiditis (HT) to Graves' disease (GD) is uncommon. Distinct immune paradigms, paucity of functioning tissue in long-standing HT, and infrequent conversion of blocking (TBAb) to stimulating (TSAb) thyrotrophin receptor antibody (TRAb) may account for this. Molecular and crystal structure analysis helps delineate TSH receptor (TSHR)/TRAb interactions in detail. Such 'fingerprinting' helps determine the behaviour and characteristics of TRAb in longitudinal studies. PATIENT: An 80-year-old woman taking thyroxine for long-standing HT became hyperthyroid. This persisted despite thyroxine withdrawal - free T3 was 7·3 pmol/l (2·6-5·7) and TSH < 0·01 mU/l (0·2-4·5) and TRAb highly positive. She had a goitre (ultrasound - HT), pretibial myxoedema, with mild inactive Graves' orbitopathy. She had RAI treatment and is on thyroxine replacement. MEASUREMENTS AND RESULTS: Blood samples at presentation (A) and 1 year (B) showed high TSAb and TPOAb activity but no TBAb. Experiments involving TSHR mutations confirmed that (i) TRAb had stable characteristics over 1 year; (ii) TSHR mutation R255D caused complete inhibition and (iii) R109A caused marked reduction of cAMP production by M22 (TSHR-stimulating human monoclonal antibody) and A and B; (iv) mutations R80A, E107A and K129A while affecting M22 had little effect on A and B. CONCLUSIONS: The reasons for an immunological paradigm shift in this elderly woman remain speculative. We believe that de-novo TSAb synthesis occurred converting her long-standing HT to GD although the mechanisms responsible remain unexplained. TRAb analysis confirmed stable autoantibody characteristics over 1 year and variable effects of TSHR mutations on TRAb and M22 function.
Assuntos
Doença de Graves/etiologia , Doença de Graves/imunologia , Doença de Hashimoto/complicações , Doença de Hashimoto/imunologia , Imunoglobulinas Estimuladoras da Glândula Tireoide/sangue , Dermatoses da Perna/etiologia , Dermatoses da Perna/imunologia , Mixedema/etiologia , Mixedema/imunologia , Idoso , Idoso de 80 Anos ou mais , Animais , Anticorpos Bloqueadores/sangue , Células CHO , Cricetinae , Cricetulus , Feminino , Doença de Graves/genética , Doença de Hashimoto/tratamento farmacológico , Humanos , Mutação , Receptores da Tireotropina/química , Receptores da Tireotropina/genética , Receptores da Tireotropina/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Tiroxina/administração & dosagem , Fatores de TempoRESUMO
Studies on the TSH receptor (TSHR) have numerous practical applications in vitro and in vivo. For example human monoclonal autoantibodies (MAbs) to the TSHR are useful reagents for in vitro diagnostics. Measurement of TSHR autoantibodies (TRAbs) is helpful in diagnosis and management of autoimmune thyroid disease. Currently available highly sensitive and specific assays to measure TRAbs use the human TSHR MAb M22 instead of the TSH. Furthermore, preparations of the human TSHR MAb M22 are useful as the World Health Organisation International Standard for thyroid stimulating antibody and for calibration of the assays for measuring TRAbs. Preparations of thermostabilised TSHR extracellular domain have recently become available and this is likely to have an impact on improvements in specificity testing for TRAb assays. In addition the stable TSHR preparations have practical application for specific immunoadsorption of patient serum TRAbs. Human TSHR MAbs also have promising prospects as new therapeutics. Autoantibodies with TSHR antagonistic activities are "natural" inhibitors of TSHR stimulation and are expected to be helpful in controlling TSHR activity in patients with Graves' disease, Graves' ophthalmopathy and thyroid cancer.
Assuntos
Doença de Graves , Receptores da Tireotropina , Anticorpos Monoclonais , Autoanticorpos , Humanos , Imunoglobulinas Estimuladoras da Glândula TireoideRESUMO
B-cells influence T-cell reactivity by facilitating antigen presentation, but the role of autoantibody-secreting B-cells in regulating T-cell responses in Type 1 diabetes is poorly defined. The aims of this study were to characterise epitopes on the IA-2 autoantigen for three monoclonal antibodies from diabetic patients by amino acid substitutions of selected residues of IA-2, establish contributions of these epitopes to binding of serum antibodies in Type 1 diabetes and relate B- and T-cell responses to overlapping determinants on IA-2. The monoclonal antibodies recognised overlapping epitopes, with residues within the 831-860 region of IA-2 contributing to binding; substitution of Glu836 inhibited binding of all three antibodies. Monoclonal antibody Fab fragments and substitution of residues within the 831-836 region blocked serum antibody binding to an IA-2 643-937 construct. IL-10-secreting T-cells responding to peptides within the 831-860 region were detected by cytokine-specific ELISPOT in diabetic patients and responses to 841-860 peptide were associated with antibodies to the region of IA-2 recognised by the monoclonal antibodies. The study identifies a region of IA-2 frequently recognised by antibodies in Type 1 diabetes and demonstrates that these responses are associated with T-cells secreting IL-10 in response to a neighbouring determinant.
Assuntos
Autoanticorpos/imunologia , Autoantígenos/imunologia , Diabetes Mellitus Tipo 1/imunologia , Epitopos de Linfócito T/imunologia , Proteínas Tirosina Fosfatases Classe 8 Semelhantes a Receptores/imunologia , Linfócitos T/imunologia , Adolescente , Adulto , Substituição de Aminoácidos , Anticorpos Monoclonais/imunologia , Criança , Epitopos de Linfócito T/genética , Feminino , Humanos , Lactente , Interferon gama/biossíntese , Interferon gama/imunologia , Interleucina-10/biossíntese , Interleucina-10/imunologia , Masculino , Proteínas Tirosina Fosfatases Classe 8 Semelhantes a Receptores/genética , Linfócitos T/metabolismo , Adulto JovemRESUMO
TSH receptor (TSHR) autoantibodies (TRAbs)activate the TSHR cyclic AMP cascade (stimulating TRAbs) or act as TSHR antagonist (blocking TRAbs), and both types inhibit TSH binding to the TSHR. Isolation of human monoclonal TSHR autoantibodies (stimulating M22 and blocking 5C9) has been a key milestone in studies of the TSHR and TSHR autoimmunity. Comparison of M22 and TSH interactions with the TSHR at the atomic level reveal that M22 heavy and light chains mimic TSH alpha and beta chains, respectively, in the way they bind to the receptor, but the evolutionary forces which have caused this close molecular mimicry are as yet completely unknown. More recently two more human monoclonal antibodies to the TSHR (K1-18 with stimulating and K1-70 with blocking activities) have been isolated from a single blood sample collected from a patient with hypothyroidism who previously presented with hyperthyroidism. K1-18 and K1-70 were derived from different lymphocytes as shown by V region genes analysis. This provides, for the first time, clear proof that a patient can produce both blocking and stimulating TRAbs at the same time. Although it has been postulated that stimulating and blocking TRAbs bind to different regions on the TSHR, our studies showed that antibodies of both types bind well to the TSHR containing only N-terminal amino acids 22-260. Whether TRAbs make contact with other parts of the TSHR in order to produce their biological effects (stimulation or blocking) remains to be elucidated.
Assuntos
Autoanticorpos/imunologia , Doenças Autoimunes/imunologia , Receptores da Tireotropina/imunologia , Doenças da Glândula Tireoide/imunologia , Animais , Anticorpos Monoclonais/química , Anticorpos Monoclonais/imunologia , Reações Antígeno-Anticorpo , Autoanticorpos/química , Humanos , Receptores da Tireotropina/química , Tireotropina/imunologiaRESUMO
This review considers recent developments in our understanding of the properties of TRAb, particularly measurement of the antibodies and their sites of action and synthesis. Two new assay methods have allowed considerable improvements in the sensitivity, specificity, precision, and ease of measuring TRAb. In particular: 1) receptor assays based on inhibition of receptor-purified labeled TSH binding to detergent-solubilized TSH receptors and 2) bioassays based on stimulation of cAMP release from monolayer cultures of isolated thyroid cells. Detailed studies with the two assays indicate that TSH receptor antibodies nearly always act as TSH agonists in patients with a history of Graves' hyperthyroidism. Studies in areas of dietary iodine sufficiency suggest that measurement of the antibodies at various stages in the course of treating Graves' disease can be of value in predicting the outcome of therapy. However, in areas of iodine deficiency, difficulties in the ability of patients' thyroid tissue to recover from the effects of antithyroid drugs may prevent the receptor antibodies from causing a relapse of thyrotoxicosis. Consequently, the predictive value of receptor antibody measurements would be expected to be lower in these geographical areas. Although patients with a history of Graves' hyperthyroidism nearly always have TRAb which act as TSH agonists, about 20% of patients with frank hypothyroidism due to autoimmune destruction of the thyroid have TRAb which act as TSH antagonists (blocking antibodies). There is some evidence that these blocking antibodies can cause hypothyroidism particularly in the neonate. With regard to the site of synthesis of TRAb, there is now direct evidence that they are synthesized by thyroid lymphocytes, particularly the lymphocytes in close proximity to thyroid follicular cells. This is consistent with the well established effects of antithyroid treatment (drugs, radioiodine, or surgery) on TRAb levels in addition to their effects on thyroid hormone synthesis. Recent studies using affinity labeling with 125I-labeled TSH have enabled elucidation of the structure of the TSH receptor. TSH receptors in human, porcine, and guinea pig thyroid tissue have a two-chain structure in which the TSH binding site is formed on the outside surface of the cell membrane by a water-soluble A subunit (Mr approximately 50 K). The A subunit is linked by a disulfide bridge and weak noncovalent bonds to the amphiphilic B subunit (Mr approximately 30 K). This subunit, which penetrates the lipid bilayer, probably forms the site for interaction of the receptor with the regulatory subunits of adenylate cyclase.(ABSTRACT TRUNCATED AT 400 WORDS)
Assuntos
Autoanticorpos/análise , Receptores da Tireotropina/imunologia , Reações Antígeno-Anticorpo , Doença de Graves/imunologia , Humanos , Conformação Proteica , Receptores da Tireotropina/análise , Tireoidite Autoimune/imunologiaRESUMO
OBJECTIVE: The glycoprotein hormones luteinizing hormone (LH), follicle-stimulating hormone (FSH), and thyrotropin (TSH) show low-level cross-reactivity between their respective receptors (R). Patient serum autoantibodies to the thyrotropin receptor (TSHR) do not appear to cross-react with the luteinizing hormone receptor (LHR) or follicle-stimulating hormone receptor (FSHR), although the concentrations of autoantibody with which it is feasible to carry out experiments of this type are limited. Consequently, we have studied the effects of high doses of the thyroid-stimulating human monoclonal autoantibody (M22) on the LHR and FSHR. DESIGN: Chinese Hamster ovary (CHO) cells stably expressing the TSHR, LHR, and FSHR and purified M22 IgG preparations were used in the study. METHODS: CHO-TSHR, CHO-LHR, and CHO-FSHR cells were incubated with bovine TSH (0.1-25mU/mL), human recombinant chorionic gonadotropin (hCG; 0.5-10mU/mL) or human recombinant FSH (100-5000mU/mL) or with M22 IgG (0.001-5.0 microg/mL), and the extracellular cyclic AMP was measured by radioimmunoassay. RESULTS: Cyclic AMP levels increased in a dose-dependent manner after incubation of CHO-TSHR cells with TSH or M22 IgG, and on a molar basis the effects of TSH and M22 were similar. Cyclic AMP stimulation was not detectable in CHO-LHR and CHO-FSHR cells after incubation with M22 IgG, whereas incubation with hCG or FSH, respectively, caused dose-dependent cyclic AMP stimulation. On a molar basis, concentrations of M22 IgG approximately 100x those of FSH causing clear stimulation were ineffective with CHO-FSHR cells. Similarly, molar concentration of M22 IgG 20,000x those of hCG causing clear stimulation had no effect on CHO-LHR cells. CONCLUSIONS: This study shows that at relatively high concentrations, M22 IgG is unable to stimulate cyclic AMP levels in CHO-LHR or CHO-FSHR cells, suggesting that TSHR autoantibodies have greater specificity for the TSHR than TSH itself.
Assuntos
Anticorpos Monoclonais/imunologia , Autoanticorpos/imunologia , Receptores do FSH/imunologia , Receptores do LH/imunologia , Animais , Anticorpos Monoclonais/metabolismo , Anticorpos Monoclonais/farmacologia , Afinidade de Anticorpos , Especificidade de Anticorpos , Autoanticorpos/metabolismo , Autoanticorpos/farmacologia , Células CHO , Gonadotropina Coriônica/farmacologia , Cricetinae , Cricetulus , Reações Cruzadas , AMP Cíclico/farmacologia , Relação Dose-Resposta Imunológica , Hormônio Foliculoestimulante/farmacologia , Expressão Gênica , Humanos , Imunoglobulinas Estimuladoras da Glândula Tireoide , Ligação Proteica/imunologia , Receptores do FSH/genética , Receptores do FSH/metabolismo , Receptores do LH/genética , Receptores do LH/metabolismo , Tireotropina/farmacologiaRESUMO
AIMS: The prevalence of islet cell, thyroid, adrenal and celiac disease related autoantibodies in patients with Type 1 diabetes mellitus (Type 1 DM) from Sri Lanka is described. DESIGN AND METHODS: Autoantibodies to glutamic acid decarboxylase 65 (GAD65Ab), protein tyrosine phosphatase IA-2 (IA-2Ab), insulin (IAAb), thyroglobulin (TgAb), thyroid peroxidase (TPOAb), TSH receptor (TRAb), 21-hydroxylase (21-OHAb) and tissue transglutaminase (tTGAb) were measured in 122 Type 1 DM patients who had low C-peptide activity or were >20 yr old at the time of diagnosis and in 100 non-diabetic blood donors. RESULTS: GAD65Ab and/or IA-2Ab were present in 74/122 (60.7%) Type 1 DM subjects with a significantly higher prevalence compared to non-diabetic controls (no. 100) (GAD65Ab-59 vs 4%; IA-2Ab-14 vs 0%; respectively) (p<0.001). The median (inter-quartile range) Type 1 DM duration in antibody positive subjects was 3.3 (0.99-6.9) vs 4.9 (1.7-7.5) yr in antibody negative subjects (p=0.23). IA-2Ab prevalence decreased with disease duration > or =5 yr (19 vs 4%) (p<0.001). There was no difference in the prevalence of TgAb (25 vs 33%)(p=0.21) and TPOAb (22 vs 18%) (p=0.48) in Type 1 DM and non-diabetic subjects. Also, there was no difference in TgAb and TPOAb prevalence in antibody positive Type 1 DM (34.7%) compared to antibody negative Type 1 DM (24.4%) subjects (p=0.24). tTGAb (3/119) and TRAb (1/119) were found in low prevalence and 21-OHAb were not detected. CONCLUSIONS: Diabetes associated autoantibodies were detected in the majority of Type 1 DM subjects, suggesting a major role for autoimmunity in the pathogenesis of Type 1 DM in Sri Lankans. The prevalence of TgAb and TPOAb in Type 1 DM subjects and non-diabetic controls was relatively high and similar in both groups.
Assuntos
Glândulas Suprarrenais/imunologia , Autoanticorpos/análise , Doença Celíaca/imunologia , Diabetes Mellitus Tipo 1/imunologia , Ilhotas Pancreáticas/imunologia , Glândula Tireoide/imunologia , Adolescente , Adulto , Idade de Início , Doença Celíaca/epidemiologia , Criança , Diabetes Mellitus Tipo 1/epidemiologia , Feminino , Glutamato Descarboxilase/imunologia , Humanos , Imunoglobulinas Estimuladoras da Glândula Tireoide , Anticorpos Anti-Insulina/análise , Iodeto Peroxidase/imunologia , Isoenzimas/imunologia , Masculino , Prevalência , Proteína Tirosina Fosfatase não Receptora Tipo 1 , Proteínas Tirosina Fosfatases/imunologia , Sri Lanka/epidemiologia , Esteroide 21-Hidroxilase/imunologia , Transglutaminases/imunologiaRESUMO
A monoclonal autoantibody (MAb) with powerful thyroid stimulating activity has been produced from lymphocytes from a patient with Graves' disease. The autoantibody and its Fab fragment bind to the thyroid stimulating hormone (TSH) receptor (TSHR) with high affinity, inhibit labelled TSH binding to the receptor and stimulate cyclic AMP production in Chinese hamster ovary cells transfected with TSHR. TSHR autoantibodies with TSH agonist or antagonist activities from patients' serum samples are effective inhibitors of labelled monoclonal autoantibody binding to TSHR. Thus, the human monoclonal autoantibody has all the characteristics of serum TSHR autoantibodies. Its availability has important implications for new studies on the pathogenesis of Graves' disease.
Assuntos
Anticorpos Monoclonais/imunologia , Autoanticorpos/imunologia , Receptores da Tireotropina/imunologia , Humanos , Fragmentos Fab das Imunoglobulinas/imunologia , Imunoglobulina G/imunologiaRESUMO
OBJECTIVE: To study possible mechanisms for the inhibition of cytochrome P450 C21 (steroid 21-hydroxylase) enzyme activity by P450 C21 autoantibodies (Abs) in vitro. DESIGN: Two possible mechanisms for the inhibition of P450 C21 enzyme activity by P450 C21 Abs were studied: (a) conformational changes in the P450 C21 molecule induced by Ab binding and (b) the effects of Ab binding to P450 C21 on the electron transfer from the nicotinamide adenine dinucleotide phosphate reduced (NADPH) cytochrome P450 reductase (CPR) to P450 C21. METHODS: The effect of P450 C21 Ab binding on the conformation of recombinant P450 C21 in yeast microsomes was studied using an analysis of the dithionite-reduced CO difference spectra. The effect of P450 C21 Abs on electron transfer was assessed by analysis of reduction of P450 C21 in the microsomes in the presence of CO after addition of NADPH. RESULTS: Our studies confirmed the inhibiting effect of P450 C21 Abs on P450 C21 enzyme activity. Binding of the Abs did not induce significant change in the P450 C21 peak at 450nm (native form) and did not produce a detectable peak at 420 nm (denatured form) in the dithionite-reduced CO difference spectra. This indicated that conformation of P450 C21 around the heme was not altered compared with the native structure. However, incubation of the P450 C21 in yeast microsomes with P450 C21 Ab inhibited the fast phase electron transfer from the CPR to P450 C21. CONCLUSIONS: Our observations suggested that the mechanism by which P450 C21 Abs inhibit P450 C21 enzyme activity most likely involves inhibition of the interaction between the CPR and P450 C21.
Assuntos
Doença de Addison/enzimologia , Autoanticorpos/metabolismo , Esteroide 21-Hidroxilase/antagonistas & inibidores , Doença de Addison/imunologia , Monóxido de Carbono/farmacologia , Ditionita , Humanos , Imunoglobulina G/metabolismo , Microssomos/metabolismo , NADPH-Ferri-Hemoproteína Redutase/metabolismo , Oxirredução , Conformação Proteica , Proteínas Recombinantes/antagonistas & inibidores , Proteínas Recombinantes/química , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/metabolismo , Espectrofotometria Ultravioleta/métodos , Esteroide 21-Hidroxilase/química , Esteroide 21-Hidroxilase/imunologia , Esteroide 21-Hidroxilase/metabolismoRESUMO
Analysis of nine mouse monoclonal antibodies (mAbs) to the thyrotropin receptor (TSHR) with TSH antagonist activity showed that only one of the mAbs (RSR B2) was an effective antagonist of the human thyroid stimulating autoantibody M22. Crystals of B2 Fab were analyzed by x-ray diffraction and a crystal structure at 3.3 A resolution was obtained. The surface charge and topography of the B2 antigen binding site were markedly different from those of the thyroid-stimulating mAb M22 and these differences might contribute to the different properties of the two mAbs. B2 (but not other mouse TSHR-specific mAbs) was also an effective antagonist of thyroid stimulating autoantibody activity in 14 of 14 different sera from patients with Graves' disease. 125I-labeled B2 bound to the TSHR with high affinity (2 x 10(10) L/mol) and patient serum TSHR autoantibodies inhibited labeled B2 binding to the receptor in a similar way to inhibition of labeled TSH binding (r = 0.75; n = 20). Furthermore, labeled B2 binding was inhibited by patient serum TSHR autoantibodies with TSH antagonist activity and also by mouse and human thyroid stimulating mAbs. Overall, mAb B2 is a powerful antagonist of thyroid stimulating autoantibodies (and TSH) thus resembling closely patient serum TSH antagonist TSHR autoantibodies. Furthermore, B2 might have potentially important in vivo applications when tissues containing the TSHR (including those in the orbit) need to be made unresponsive to stimulating autoantibodies.
Assuntos
Anticorpos Monoclonais/imunologia , Autoanticorpos/imunologia , Doença de Graves/imunologia , Receptores da Tireotropina/imunologia , Tireotropina/imunologia , Animais , Anticorpos Monoclonais/química , Células CHO , Cricetinae , Cristalografia por Raios X , Doença de Graves/terapia , Humanos , Fragmentos Fab das Imunoglobulinas/química , Fragmentos Fab das Imunoglobulinas/imunologia , Radioisótopos do Iodo , Camundongos , Estrutura Terciária de Proteína , Receptores da Tireotropina/metabolismo , Tireotropina/metabolismoRESUMO
Human 21-hydroxylase (21-OH) genes containing various mutations, truncations, and deletions were expressed in yeast, and autoantibody binding was studied by Western blotting using patient sera and rabbit antibodies to 21-OH. 21-OH autoantibodies in 13 Addisonian sera showed a marked reduction in their ability to recognize 21-OH mutated at Pro453-->Ser (mean +/- SD, 31 +/- 9% of binding to wild type), whereas the effect on rabbit antibody binding was small (88 +/- 11% of binding to wild type; n = 7). Mutation at Arg339-->His had a less pronounced effect on autoantibody binding (85 +/- 11% of binding to wild type; n = 13) and caused a small enhancement of rabbit antibody binding (124 +/- 16% of binding to wild type; n = 7). These studies indicate that Pro453 has a key role in forming an autoantigenic epitope on 21-OH. It is important to note, however, that the Pro453 mutation caused only partial loss of autoantibody binding, i.e. all Addisonian sera studied still reacted with the mutated protein. This may indicate that each serum sample contains at least two different populations of 21-OH autoantibodies, only one of which recognizes a site dependent on Pro453. A series of more extensive modifications of the 21-OH sequence, including truncations (amino acids 460-494, 448-494, and 418-494) and deletions (amino acids 165-379, 142-240, and 142-280) indicated that most of the sequence of amino acids from 241-494 is important for autoantibody binding. The involvement of such an extensive region of the molecule suggests that the binding sites are generated by three-dimensional folding, with Pro453 having a critical role in forming at least one major autoantigenic epitope.
Assuntos
Glândulas Suprarrenais/imunologia , Autoanticorpos/metabolismo , Mutação , Esteroide 21-Hidroxilase/genética , Esteroide 21-Hidroxilase/imunologia , Autoantígenos/química , Autoantígenos/imunologia , Sequência de Bases , Western Blotting , Deleção de Genes , Expressão Gênica , Humanos , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Mapeamento por Restrição , Saccharomyces cerevisiae/genética , Esteroide 21-Hidroxilase/química , Relação Estrutura-AtividadeRESUMO
Autoantibodies to steroid 21-hydroxylase (21-OH) are characteristic of adult onset Addison's disease and we have investigated the effects of these autoantibodies on recombinant human 21-OH enzyme activity. Antibody preparations from 11/11 Addison sera inhibited the ability of 21-OH to convert progesterone to deoxycorticosterone with 8 IgGs showing almost complete inhibition, 2 partial inhibition and 1 weak inhibition. Control IgGs from patients with autoimmune thyroid disease and normal blood donors had little or no effect on 21-OH activity. Our results suggest that 21-OH autoantibodies have the potential to contribute to adrenal failure in Addison's disease by inhibiting the 21-OH enzyme.
Assuntos
Doença de Addison/etiologia , Insuficiência Adrenal/imunologia , Autoanticorpos/fisiologia , Doenças Autoimunes/etiologia , Esteroide 21-Hidroxilase/imunologia , Esteroide 21-Hidroxilase/fisiologia , Doença de Addison/imunologia , Adulto , Autoanticorpos/análise , Autoanticorpos/imunologia , Doenças Autoimunes/imunologia , Humanos , Imunoglobulina G/análiseRESUMO
Adrenal autoantibodies (ACA) are markers of adrenal cortex involvement in idiopathic Addison's disease. Recently the 21-hydroxylase (21-OH) enzyme has been discovered to be the major autoantigen of the ACA. A potential role of these antibodies in determining adrenal failure by inhibition of the 21-OH has been recently postulated. To test this hypothesis, cortisol and aldosterone (final products of adrenal steroid synthesis) and 17-hydroxyprogesterone (17-OH-progesterone) (as a marker of 21-OH impairment) have been investigated in baseline conditions and after ACTH (1-24) stimulation test in a group of 42 patients positive for both ACA and 21-OH autoantibodies. Patients were divided into five groups according to the stages (0-4) of adrenal failure. With progression toward overt Addison's disease, baseline 17-OH-progesterone, cortisol, and aldosterone remained almost unchanged but with impairment of their responses to ACTH (1-24) stimulation. The 17-OH-progesterone/cortisol ration remained normal both in basal conditions and after stimulation at stages 0-3. At stage 4 (overt Addison's disease), this ratio increased in baseline condition with no changes after ACTH (1-24), probably because of persistent 17-OH-progesterone gonadal production. In conclusion, there was a progressive and concomitant impairment of the synthesis of all steroids tested over various phases of adrenal failure. The pattern of response of the 17-OH-progesterone/cortisol ratio to ACTH stimulation in patients with 21-OH autoantibodies was not consistent with the autoantibodies inhibiting the 21-OH activity. This suggests that the inhibiting effect of 21-OH autoantibodies on 21-OH activity is not usually evident in vivo.
Assuntos
Doenças das Glândulas Suprarrenais/sangue , Doenças Autoimunes/sangue , Hormônios/sangue , Esteroide 21-Hidroxilase/antagonistas & inibidores , Doenças das Glândulas Suprarrenais/fisiopatologia , Glândulas Suprarrenais/imunologia , Hormônio Adrenocorticotrópico/sangue , Adulto , Autoanticorpos/análise , Doenças Autoimunes/fisiopatologia , Progressão da Doença , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Renina/sangueRESUMO
An investigation of the sugar groups on recombinant human TSH receptors (TSHR) expressed in CHO-K1 cells and solubilized with detergents is described. Western blotting studies with TSHR monoclonal antibodies showed that the receptor was present principally as two bands with approximate molecular masses of 120 and 100 kDa. Further blotting studies using lectins and/or involving treatment with different glycosidases indicated that the 100-kDa band contained about 16 kDa of high mannose-type sugars, and the 120-kDa band contained about 33 kDa of complex-type sugars. It was possible to separate the 120- and 100-kDa components of the TSHRs by lectin affinity chromatography. In particular, Galanthus nivalis lectin, which binds high mannose-type sugars, bound the 100-kDa band, but not the 120-kDa band, whereas Datura stramonium lectin, which binds complex-type sugars, bound the 120-kDa band, but not the 100-kDa band. 125I-Labeled TSH binding studies with the various lectin column fractions showed that TSH-binding activity was principally associated with the complex-type sugar containing the 120-kDa form of the receptor rather than the high mannose-containing 100-kDa form. During peptide chain glycosylation, high mannose-type sugar residues are attached first and then modified by the formation of complex type structures to form the mature glycoprotein. Our data suggest that in the case of the TSH receptor, this type of posttranslational processing has an important role in forming the TSH-binding site.